Gene transfer to airway epithelia could provide an important new treatment for cystic fibrosis (CF) lung disease. A common problem with current vector systems is that the efficiency of gene transfer to differentiated human airway epithelia is limited. In work supported by this Program, we have investigated the advantages and limitations of adenoviral and non-viral vectors. By combining the two systems, we have utilized their unique advantages and avoided many of the limitations. In so doing, we have developed novel vector systems, including Ad:CaPi co- precipitates. This vector shows markedly enhanced gene transfer to differentiated airway epithelia. Moreover, preliminary data suggest that the Ad:CaPi co-precipitates do not produce additional toxicity. In this Project we focus on six questions. 1) How do Ad:CaP co-precipitates infect cells? 2) What properties of Ad:CaPi co-precipitates are important for gene transfer? 3) What cells are targeted by Ad:CaPi co- precipitates and other complexes? 4) Can complexes shield adenovirus from neutralizing antibodies? 5) Can other vectors, including AAV, be incorporated into CaPi co-precipitates? The results of these studies will improve our understanding of the mechanisms and barriers and gene transfer, will have application of several different vector systems, and should ultimately lead to improved gene transfer for CF airway disease.